Outer Tissue Research Articles

Glutathione accelerates the cell cycle and cellular reprogramming in plant regeneration.

The plasticity of plant cells underlies their wide capacity to regenerate, with increasing evidence in plants and animals implicating cell cycle dynamics in cellular reprogramming. To investigate the cell cycle during cellular reprogramming, we developed a comprehensive set of cell cycle phase markers in the Arabidopsis root. Using single-cell RNA-seq profiles and live imaging during regeneration, we found that a subset of cells near an ablation injury dramatically increases division rate by truncating G1. Cells in G1 undergo a transient nuclear peak of glutathione (GSH) prior to coordinated entry into S phase followed by rapid divisions and cellular reprogramming. A symplastic block of the ground tissue impairs regeneration, which is rescued by exogenous GSH. We propose a model in which GSH from the outer tissues is released upon injury licensing an exit from G1 near the wound to induce rapid cell division and reprogramming.

The Management of Root Perforation: A Review of the Literature.

Root canal perforation is defined as an opening or communication of the root to the outer oral tissue that occurs either accidentally during root canal therapy or due to unknown pathological causes, which generally result in complications in patientsdue to the passage of microorganisms to the surrounding tissues. These complications may necessitate extraction of the affected tooth. In this literature review, 42 articles were reviewed initially; 33 of them were included in the final analysis based on exclusion and inclusion criteria. Root perforation may occur due to caries, external and internal resorption, trauma, as well as iatrogenic causes. Many studies have suggested that perforation can be identified by radiographs, unexpected bleeding and pain during instrumentation, blood on paper points, electronic apex locator, and dental operating microscope. Several materials have been used in endodontic perforation repairs, such as indium foil, super 2-ethoxybenzoic acid (super EBA), as well as bioceramic materials like mineral trioxide aggregate (MTA) and biodentine. These materials must achieve biocompatibility, bacteriostatic ability, and radio-opacity. Many studies have reported the superior properties of MTA in comparison with other materials. Based on our literature review, management of root perforation should be performed based on multiple significant factors. Clinicians must possess a comprehensive understanding of tooth anatomy to prevent perforation occurrence. Furthermore, clinical management and prognosis of root perforation have been enhanced by the availability of advanced materials and sealing techniques.

Guardians of the clams: a decadal monitoring effort of endangered giant clams by citizen scientists

ABSTRACT Giant clams are highly conspicuous marine bivalves in the tropical oceans because of their large shells and colourful outer mantle tissues. This makes them an appropriate iconic species for both trained and untrained citizen science surveys. In Singapore, because of good public awareness of giant clams, they are often reported by citizen scientists, who document marine species using photography. Furthermore, these opportunistic records are captured in a comprehensive public blog (Wild Shores of Singapore; https://wildshores.blogspot.com) with specific details of the field surveys. This study presents the first feasible effort in establishing a long-term dataset based on citizen science observations to report giant clam species-level population status and trends over time. Between 2011 and 2020, citizen scientists covered 8.53 km2 of intertidal reef areas and consistently recorded only two giant clam species (Tridacna squamosa and T. crocea) annually. Despite the more comprehensive coverage and sampling of reef areas, the citizen scientists did not encounter the other native species: Hippopus hippopus, T. gigas and T. maxima. Also, the respective densities of T. squamosa and T. crocea were 0.00120 and 0.00034 individuals per 100 m2, which makes them the smallest known giant clam populations in the Indo-Pacific region. Regardless, the sampling efforts of citizen scientists were generally similar across the years, but future surveys could adapt different survey protocols for the respective giant clam species. Findings from this study also support the marine conservation efforts in Singapore, such as using active intervention measures to protect their giant clams. Additionally, lessons learnt from these citizen science observations can be harnessed to encourage monitoring of giant clams throughout the Indo-Pacific region.

In Vivo Lifetime Imaging of the Internal O2 Dynamics in Corals with near-Infrared-Emitting Sensor Nanoparticles.

Mapping of O2 with luminescent sensors within intact animals is challenging due to attenuation of excitation and emission light caused by tissue absorption and scattering as well as interfering background fluorescence. Here we show the application of luminescent O2 sensor nanoparticles (∼50-70 nm) composed of the O2 indicator platinum(II) tetra(4-fluoro)phenyltetrabenzoporphyrin (PtTPTBPF) immobilized in poly(methyl methacrylate-co-methacrylic acid) (PMMA-MA). We injected the sensor nanoparticles into the gastrovascular system of intact colony fractions of reef-building tropical corals that harbor photosynthetic microalgae in their tissues. The sensor nanoparticles are excited by red LED light (617 nm) and emit in the near-infrared (780 nm), which enhances the transmission of excitation and emission light through biological materials. This enabled us to map the internal O2 concentration via time-domain luminescence lifetime imaging through the outer tissue layers across several coral polyps in flowing seawater. After injection, nanoparticles dispersed within the coral tissue for several hours. While luminescence intensity imaging showed some local aggregation of sensor particles, lifetime imaging showed a more homogeneous O2 distribution across a larger area of the coral colony. Local stimulation of symbiont photosynthesis in corals induced oxygenation of illuminated tissue areas and formation of lateral O2 gradients toward surrounding respiring tissues, which were dissipated rapidly after the onset of darkness. Such measurements are key to improving our understanding of how corals regulate their internal chemical microenvironment and metabolic activity, and how they are affected by environmental stress such as ocean warming, acidification, and deoxygenation. Our experimental approach can also be adapted for in vivo O2 imaging in other natural systems such as biofilms, plant and animal tissues, as well as in organoids and other cell constructs, where imaging internal O2 conditions are relevant and challenging due to high optical density and background fluorescence.

Nearly instantaneous stem diameter response to fluctuations in the atmospheric water demand.

Changes in vapour pressure deficit can lead to the depletion and replenishment of stem water pools to buffer water potential variations in the xylem. Yet, the precise velocity at which stem water pools track environmental cues remains poorly explored. Nine eucalyptus seedlings grown in a glasshouse experienced high-frequency environmental oscillations and their stem radial variations (ΔR) were monitored at a 30-s temporal resolution in upper and lower stem locations and on the bark and xylem. The stem ΔR response to vapour pressure deficit changes was nearly instantaneous (<1min), while temperature lagged behind stem ΔR. No temporal differences in the stem ΔR response were observed between locations. Punctual gravimetric measurements confirmed the synchrony between transpiration and stem ΔR dynamics. These results indicate (i) that stem-stored water can respond to the atmospheric evaporative demand much faster than commonly assumed and (ii) that the origin of the water released to the transpiration stream seems critical in determining time lags in stem water pool dynamics. Near-zero time lags may be explained by the high elasticity of eucalyptus woody tissues and the predominant water use from the xylem, circumventing the hydraulic radial barriers to water flow from/to the outer tissues.

The structural organization of the outer tissues in the gametophytic stem of the umbrella moss Hypnodendron menziesii optimizes load bearing.

The ultrastructural design and biochemical organization of the significantly thickened outer tissues of the gametophytic stem of Hypnodendron menziesii optimizes load bearing of the stem. Hypnodendron menziesii is a bryoid umbrella moss growing in high humid conditions on the forest floors of New Zealand. The erect gametophyte bears up to eight whorls of branches in succession, spreading across the stem that bears the heavy weight of branches with highly hydrated leaves. Our investigation using a combination of light microscopy, transmission electron microscopy (TEM), scanning electron microscopy (SEM), and TEM-immunolabeling techniques provided novel information on the structural design and biochemical organization of greatly thickened cell walls of epidermal, hypodermal, and outermost cortical tissues, comparing underlying thin-walled cortical tissues in the gametophytic stem. Probing into the ultrastructure of the cell wall architecture of these target tissues by TEM and SEM revealed the cell walls to display a multilamellar organization, in addition to demonstrating the presence of an electron-dense substance in the cell wall, presumably flavonoids. The pattern of distribution and concentration of rhamnogalacturonan, homogalacturonan, and heteromannan, as determined by immunogold labeling, suggests that it is the combination of structural and molecular design of the cell wall that may optimize the mechanical function of the epidermal, hypodermal, and outer cortical tissues. Statistical relationships between the overall thickness of epidermal, hypodermal, and outer cortical cell walls, the lumen area of cells and the percentage area of cell wall occupied in these tissues at different heights of the stem, and thickness of secondary cell wall layers (L1-L4/5) were explored. The results of these analyses unequivocally support the contribution of outer tissues to the mechanical strength of the resilient stem.

Spatial multi-omics of human skin reveals KRAS and inflammatory responses to spaceflight

Spaceflight can change metabolic, immunological, and biological homeostasis and cause skin rashes and irritation, yet the molecular basis remains unclear. To investigate the impact of short-duration spaceflight on the skin, we conducted skin biopsies on the Inspiration4 crew members before (L-44) and after (R + 1) flight. Leveraging multi-omics assays including GeoMx™ Digital Spatial Profiler, single-cell RNA/ATAC-seq, and metagenomics/metatranscriptomics, we assessed spatial gene expressions and associated microbial and immune changes across 95 skin regions in four compartments: outer epidermis, inner epidermis, outer dermis, and vasculature. Post-flight samples showed significant up-regulation of genes related to inflammation and KRAS signaling across all skin regions. These spaceflight-associated changes mapped to specific cellular responses, including altered interferon responses, DNA damage, epithelial barrier disruptions, T-cell migration, and hindered regeneration were located primarily in outer tissue compartments. We also linked epithelial disruption to microbial shifts in skin swab and immune cell activity to PBMC single-cell data from the same crew and timepoints. Our findings present the inaugural collection and examination of astronaut skin, offering insights for future space missions and response countermeasures.

Type of stomata and peel structure associated with programmed cell death of senescent spotting in banana

Senescent spotting or brown spot on bananas, is regarded as a form of programmed cell death (PCD). These spots appear in areas where stomata are present on the banana peel, there is no report describing the relationship between these spots and stomata on peel. This research aims to investigate whether senescent spotting in banana is involved in the PCD of stomata on peel. Three banana cultivars, ‘Khai-Kasetsart 2’ (KK2), ‘Dwarf Cavendish’ (CVD), and ‘Hom Thong’ (HT) bananas were examined for senescent spotting. KK2 banana exhibited more severe senescent spotting than CVD banana, while HT bananas did not display brown spot. Electrolyte leakage and DNA degradation increased before the spotting was observed in KK2 and CVD bananas. The peel structure revealed compact cells in CVD and HT bananas, whereas KK2 banana had a loose cell structure. Notably, KK2 banana had a high stomata density correlate with weight loss. Interestingly, the stomata type of KK2 banana was the same as CVD banana, being the typical type, while HT had a sunken type. The spotting zone of KK2 and CVD bananas revealed that stomata were more open than in the healthy zone. Additionally, the outer ground tissue of spotting zone flattened and collapsed, while the healthy zone did not exhibit this flattening and collapsing. Observations of cell morphology in the spotting zone of KK2 banana showed a detached plasma membrane, organelle destruction, and the presence of cell corpse with an absence of cytoplasm clearance. In conclusion, the result provides an evidence that stomata type on the banana peel involve in PCD, resulting in senescent spotting.

Anatomical limitations in adventitious root formation revealed by magnetic resonance imaging, infrared spectroscopy, and histology of rose genotypes with contrasting rooting phenotypes.

Adventitious root (AR) formation is one of the most important developmental processes in vegetative propagation. Although genotypic differences in rose rooting ability are well known, the causal factors are not well understood. The rooting of two contrasting genotypes, 'Herzogin Friederike' and 'Mariatheresia', was compared following a multiscale approach. Using magnetic resonance imaging, we non-invasively monitored the inner structure of stem cuttings during initiation and progression of AR formation for the first time. Spatially resolved Fourier-transform infrared spectroscopy characterized the chemical composition of the tissues involved in AR formation. The results were validated through light microscopy and complemented by immunolabelling. The outcome demonstrated similarity of both genotypes in root primordia formation, which did not result in root protrusion through the shoot cortex in the difficult-to-root genotype 'Mariatheresia'. The biochemical composition of the contrasting genotypes highlighted main differences in cell wall-associated components. Further spectroscopic analysis of 15 contrasting rose genotypes confirmed the biochemical differences between easy- and difficult-to-root groups. Collectively, our data indicate that it is not the lack of root primordia limiting AR formation in these rose genotypes, but the firmness of the outer stem tissue and/or cell wall modifications that pose a mechanical barrier and prevent root extension and protrusion.

The content of cannabinoids in common hemp plants in the conditions of the Republic of Bashkortostan

Cannabis sativa L. – a herbaceous plant. Recently, interest in this fast-growing plant has increased again due to its multipurpose use: It is indeed a treasure trove of phytochemicals and a rich source of both cellulose and wood fibers. The pharmaceutical and construction sectors are equally interested in this plant, since its metabolites have powerful biological activity for human health, and stem outer and inner tissues can be used to manufacture bioplastics and concrete-like materials, respectively. Cannabinoids are the most studied group of compounds, mainly because of their wide range of pharmaceutical effects on the human body, including psychotropic activity. The content of the main cannabinoids in common hemp plants of the Vera, Nadezhda, Surskaya, Omegadar-1 varieties for 2020-2023 has been studied. The predominant cannabinoid in all varieties during the years of the study is CBD. The studied common hemp varieties in the years of research fully comply with the requirements of the law. The THC content did not exceed 0.1%.

Emerging therapies for geographic atrophy

Geographic atrophy (GA) is a late‐stage form of age‐related macular degeneration (AMD) characterized by progressive degeneration of photoreceptors, retinal pigment epithelium (RPE), and choriocapillaris primarily in the macula. It was shown in AREDS studies that supplementation of high‐dose antioxidant vitamins (C, E, and beta‐carotene) and zinc, decrease the risk of progression to advanced AMD.It was suggested that the complement system, a part of the human immune system, plays an important role in the GA pathophysiology. Different complement genes have been linked to AMD in genome‐wide association studies, and the odds of GA were estimated to be 2.5 times higher per copy of a common risk allele (Y402H) of the CFH gene in individuals of European ancestry. In histologic studies complement proteins were found in drusen as well as elevated levels of complement proteins in the outer retinal tissue of postmortem eyes with AMD. Moreover, higher levels of activated complement products in plasma have been found in GA patients. Thus, several treatments targeting the complement system are investigated. The drugs that showed phase 3 efficacy and safety include pegcetacoplan and avacincaptad pegol, which are complement C3 and complement C5 inhibitors, respectively.

Optical coherence tomography angiography of choroidal neovascularization in long-chain 3-hydroxyacyl-CoA dehydrogenase deficiency (LCHADD)

PurposeTo report the clinical utility of optical coherence tomography angiography (OCTA) for demonstrating choroidal neovascularization (CNV) associated with Long-Chain 3-Hydroxyacyl-CoA Dehydrogenase Deficiency (LCHADD) retinopathy. MethodsThirty-three participants with LCHADD (age 7–36 years; median 17) were imaged with OCTA and the Center for Ophthalmic Optics & Lasers Angiography Reading Toolkit (COOL-ART) software was implemented to process OCTA scans. ResultsSeven participants (21 %; age 17–36 years; median 25) with LCHADD retinopathy demonstrated evidence of CNV by retinal examination or presence of CNV within outer retinal tissue on OCTA scans covering 3 × 3 and/or 6 × 6-mm. These sub-clinical CNVs are adjacent to hyperpigmented areas in the posterior pole. CNV presented at stage 2 or later of LCHADD retinopathy prior to the disappearance of RPE pigment in the macula. ConclusionOCTA can be applied as a non-invasive method to evaluate the retinal and choroidal microvasculature. OCTA can reveal CNV in LCHADD even when the clinical exam is inconclusive. These data suggest that the incidence of CNV is greater than expected and can occur even in the early stages of LCHADD retinopathy.

How the interplay between harvest time and climatic conditions drives the dynamics of hemp (Cannabis sativa L.) field retting

Dew retting, the selective biodegradation of industrial hemp (Cannabis sativa L.) stems after harvest, is a key field process for plant fiber use. The stage of maturity of the crop and the weather conditions during the subsequent dew retting process differ with hemp harvest time, which depends on the hemp valorization scenario. In this work, the aim was to investigate and rank the important factors that drive hemp retting, such as hemp harvest date, weather during retting, soil type under hemp mulch, and their interactions under agricultural field conditions. To this end, an experimental field trial was set up in the northeastern area of France (Champagne region), with two hemp harvest scenarios and two types of soil common to the agricultural area. Physicochemical hemp characteristics and climatic conditions were dynamically monitored during the retting periods. Dynamics of stem characteristics determined at the biomass level (dry matter loss) and the outer tissue level (surface analysis, chemical and thermal analysis) showed that the kinetics of retting followed similar patterns regardless of soil type for both harvest date scenarios. The kinetics were strongly and linearly related to air temperature, expressed as the number of normalized days at 15 °C, and to cumulative radiation during retting periods. The cumulative amounts of rain and dew explained the kinetics less than temperature and radiation, regardless of the retting scenario considered. The strong relationship observed between colorimetry and infrared spectroscopic data for characterizing the retting progress for both harvest scenarios paves the way for the development of tools to monitor changes in hemp stem quality.

Selective Extracellular Matrix Guided Mesenchymal Stem Cell Self-Aggregate Engineering for Replication of Meniscal Zonal Tissue Gradient in a Porcine Meniscectomy Model.

Degenerative meniscus tears (DMTs) are prevalent findings in osteoarthritic knees, yet current treatment is mostly limited to arthroscopic partial meniscectomy rather than regeneration, which further exacerbates arthritic changes. Translational research regarding meniscus regeneration is hindered by the complex, composite nature of the meniscus which exhibit a gradient from inner cartilage-like tissue to outer fibrous tissue, as well as engineering hurdles often requiring growth factors and cross-linking agents. Here, a meniscus zonal tissue gradient is proposed using zone-specific decellularized meniscus extracellular matrix (DMECM) and autologous synovial mesenchymal stem cells (SMSC) via self-aggregation without the use of growth factors or cross-linking agents. Combination with zone-specific DMECM during self-aggregation of MSCs forms zone-specific meniscus tissue that reflects the respective DMECM harvest site. The implantation of these constructs leads to the regeneration of meniscus tissue resembling the native meniscus, demonstrating inner cartilaginous and outer fibrous characteristics as well as recovery of native meniscal microarchitecture in a porcine partial meniscectomy model at 6 months. In all, the findings offer a potential regenerative therapy for DMTs that may improve current partial meniscectomy-based patient care.

Metabolic Responses of Normal Rat Kidneys to a High Salt Intake.

In this study, novel methods were developed, which allowed continuous (24/7) measurement of arterial blood pressure and renal blood flow in freely moving rats and the intermittent collection of arterial and renal venous blood to estimate kidney metabolic fluxes of O2 and metabolites. Specifically, the study determined the effects of a high salt (HS; 4.0% NaCl) diet upon whole kidney O2 consumption and arterial and renal venous plasma metabolomic profiles of normal Sprague-Dawley rats. A separate group of rats was studied to determine changes in the cortex and outer medulla tissue metabolomic and mRNAseq profiles before and following the switch from a 0.4% to 4.0% NaCl diet. In addition, targeted mRNA expression analysis of cortical segments was performed. Significant changes in the metabolomic and transcriptomic profiles occurred with feeding of the HS diet. A progressive increase of kidney O2 consumption was found despite a reduction in expression of most of the mRNA encoding enzymes of TCA cycle. A novel finding was the increased expression of glycolysis-related genes in Cx and isolated proximal tubular segments in response to an HS diet, consistent with increased release of pyruvate and lactate from the kidney to the renal venous blood. Data suggests that aerobic glycolysis (eg, Warburg effect) may contribute to energy production under these circumstances. The study provides evidence that kidney metabolism responds to an HS diet enabling enhanced energy production while protecting from oxidative stress and injury. Metabolomic and transcriptomic analysis of kidneys of Sprague-Dawley rats fed a high salt diet.

An Intra-Tissue Radiometry Microprobe for Measuring Radiance In Situ in Living Tissue.

Organisms appear opaque largely because their outer tissue layers are strongly scattering to incident light; strongly absorbing pigments, such as blood, typically have narrow absorbances, such that the mean free path of light outside the absorbance peaks can be quite long. As people cannot see through tissue, they generally imagine that tissues like the brain, fat, and bone contain little or no light. However, photoresponsive opsin proteins are expressed within many of these tissues, and their functions are poorly understood. Radiance internal to tissue is also important for understanding photosynthesis. For example, giant clams are strongly absorbing yet maintain a dense population of algae deep in the tissue. Light propagation through systems like sediments and biofilms can be complex, and these communities can be major contributors to ecosystem productivity. Therefore, a method for constructing optical micro-probes for measuring scalar irradiance (photon flux intersecting a point) and downwelling irradiance (photon flux crossing a plane perpendicularly) to better understand these phenomena inside living tissue has been developed. This technique is also tractable in field laboratories. These micro-probes are made from heat-pulled optical fibers that are then secured in pulled glass pipettes. To change the angular acceptance of the probe, a 10-100 µm sized sphere of UV-curable epoxy mixed with titanium dioxide is then secured to the end of a pulled, trimmed fiber. The probe is inserted into living tissue, and its position is controlled using a micromanipulator. These probes are capable of measuring in situ tissue radiance at spatial resolutions of 10-100 µm or on the scale of single cells. These probes were used to characterize the light reaching the adipose and brain cells 4 mm below the skin of a living mouse and to characterize the light reaching similar depths within living algae-rich giant clam tissue.

A redundant transcription factor network steers spatiotemporal Arabidopsis triterpene synthesis.

Plant specialized metabolites modulate developmental and ecological functions and comprise many therapeutic and other high-value compounds. However, the mechanisms determining their cell-specific expression remain unknown. Here we describe the transcriptional regulatory network that underlies cell-specific biosynthesis of triterpenes in Arabidopsis thaliana root tips. Expression of thalianol and marneral biosynthesis pathway genes depends on the phytohormone jasmonate and is limited to outer tissues. We show that this is promoted by the activity of redundant bHLH-type transcription factors from two distinct clades and coactivated by homeodomain factors. Conversely, the DOF-type transcription factor DAG1 and other regulators prevent expression of the triterpene pathway genes in inner tissues. We thus show how precise expression of triterpene biosynthesis genes is determined by a robust network of transactivators, coactivators and counteracting repressors.

Evidence for Sex-dependent Bmal1 Control of Mitochondrial Dynamics in Rat Kidney

The circadian clock regulates Na+ transport with a necessary diurnal variation for optimal health. The clock gene, Bmal1, is known to regulate mitochondrial O 2 consumption, but whether or how this occurs in the kidney has not been established. An estimated 95% of O 2 consumption in the kidney is linked to sodium transport. We developed and used a novel Bmal1 knockout rat on a Sprague Dawley background. Male Bmal1 -/- rats do not have the typical night-day difference in Na + excretion, while female rats maintain this pattern. We recently observed that ADP-dependent O 2 consumption in permeabilized kidney tissue was significantly higher in both sexes of Bmal1 -/- rats but only during the active/dark period. We also observed a significant increase in cytochrome c oxidase activity in Bmal1 -/- rats during the dark vs. light period (164 ± 40 vs. 306 ± 40 pmol/s·mg; p=0.0031, t-test). Since these effects were observed in both male and female rats, these changes in mitochondrial activity cannot explain the difference in diurnal sodium excretion. Since mitochondrial morphology can impact oxidative phosphorylation and energy availability, we hypothesized that Bmal1 regulates mitochondrial morphology in the kidney. Male and female global Bmal1 +/+ and Bmal1 -/- rats at 12-14 weeks of age were maintained in regular 12:12 LD cycles. Outer renal medullary tissue was dissected and prepared for mRNA expression and transmission electron microscopy (EM). For EM, tissues were obtained at light (ZT2-4) or dark (ZT14-16) periods to correspond with the minimum and maximum whole-body energy consumption and peak and trough Bmal1 protein expression. Digital droplet PCR mRNA expression was conducted from tissue collected at 4hr intervals over 24hrs. EM images appear to show an elongation of mitochondrial morphology in females but not male Bmal1 -/- during the active time of day. Mitofusion1 (Mfn1) and fission 1 (Fis1) are genes that regulate mitochondrial structural dynamics. Mfn1 and Fis1 mRNA expression over 24hrs showed a significant interaction between genotype and time of day in male rats (p=0.004 and p=0.020, respectively, 2way ANOVA). In contrast, there were no significant differences in overall Mfn1 or Fis1 expression between female Bmal1 +/+ and Bmal1 -/- rats. Our findings demonstrate that Bmal1 is crucial in maintaining mitochondrial respiration coupling and ATP generation in the kidney. Furthermore, we suggest that female Bmal1 -/- rats can better compensate for impaired respiration by mitochondrial morphological changes when the molecular clock is dysfunctional. We further propose that the rhythmicity of Na + excretion in females, but not male, Bmal1 -/- rats is due to adjustments in mitochondrial morphology. T32 HL007457, P01HL136267, AHA 908953 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Degeneration of muscle spindles in a murine model of Pompe disease

Pompe disease is a debilitating medical condition caused by a functional deficiency of lysosomal acid alpha-glucosidase (GAA). In addition to muscle weakness, people living with Pompe disease experience motor coordination deficits including an instable gait and posture. We reasoned that an impaired muscle spindle function might contribute to these deficiencies and therefore analyzed proprioception as well as muscle spindle structure and function in 4- and 8-month-old Gaa−/− mice. Gait analyses showed a reduced inter-limb and inter-paw coordination in Gaa−/− mice. Electrophysiological analyses of single-unit muscle spindle proprioceptive afferents revealed an impaired sensitivity of the dynamic and static component of the stretch response. Finally, a progressive degeneration of the sensory neuron and of the intrafusal fibers was detectable in Gaa−/− mice. We observed an increased abundance and size of lysosomes, a fragmentation of the inner and outer connective tissue capsule and a buildup of autophagic vacuoles in muscle spindles from 8-month-old Gaa−/− mice, indicating lysosomal defects and an impaired autophagocytosis. These results demonstrate a structural and functional degeneration of muscle spindles and an altered motor coordination in Gaa−/− mice. Similar changes could contribute to the impaired motor coordination in patients living with Pompe disease.

Sequestration of Ca in simian nasal mucosa: Determination of Ca molarity in ex vivo tissue by simultaneous off-axis Scanning Transmission Ion Microscopy, Particle Induced X-Ray Emission and Elastic Backscattering Spectrometry

A technique to determine the molar concentration of elements in ex vivo tissue sections by Particle Induced X-ray Emission (PIXE) has been developed. The method is based on simultaneous off-axis scanning transmission ion microscopy (OA-STIM) and Elastic Backscattering Spectrometry (EBS) measurement of the sample thickness and major element (H, C, N, O) composition. The method was applied to determine the molarity of localised Ca concentration hot-spots in the outer epithelium tissue of nasal mucosa of a rhesus macaque subject infected with simian immunodeficiency virus (SIV). The results show Ca sequestration in concentration hot-spots and outer epithelial tissue that significantly exceeded the Ca concentration in the surrounding tissues. This may originate from mineralisation and/or Ca enhancement in goblet cells.